1. Technical Field
The present invention relates to a method for producing L-glutamic acid by fermentation. L-glutamic acid is an important amino acid as food, medicines and so forth.
2. Related Art
L-glutamic acid has conventionally been produced mainly by fermentation utilizing L-glutamic acid-producing coryneform bacteria belonging to the genus Brevibacterium, Corynebacterium, or Microbacterium, or variants thereof (Amino Acid Fermentation, Gakkai Shuppan Center, pp.195-215, 1986).
In recent years, there have been disclosed techniques for breeding L-glutamic acid-producing coryneform bacteria by utilizing genetic recombination techniques. For example, it has been disclosed that an α-ketoglutarate dehydrogenase-deficient L-glutamic acid-producing coryneform bacterium in which the enzyme has been deleted by destroying the gene for the enzyme, for example, can produce and accumulate a marked amount of L-glutamic acid when it is cultured in a medium containing an excessive amount of biotin without addition of any biotin activity inhibiting substance such as surface active agents and penicillin to the medium (WO95/34672).
Under an aerobic condition, L-glutamic acid is biosynthesized from saccharides such as glucose. That is, pyruvate produced in the glycolysis pathway is modified with acetyl-CoA, and enters into the tricarboxylic acid cycle, and L-glutamic acid is biosynthesized from an intermediate of the tricarboxylic acid cycle, α-ketoglutarate, through a reaction catalyzed by glutamate dehydrogenase or glutamine synthetase/glutamate synthase branched off from the tricarboxylic acid cycle at the α-ketoglutarate. Based on these findings, there has been disclosed a method for producing L-glutamic acid utilizing a glutamic acid-producing coryneform bacterium harboring a recombinant DNA that contains a gene encoding glutamate dehydrogenase, isocitrate dehydrogenase, aconitate hydratase, and citrate synthase (Japanese Patent Laid-open No. 63-214189).
While the productivity of L-glutamic acid has been improved by breeding of microorganisms such as those mentioned above and by improvement of production methods therefor, it is still desired to develop techniques for further increasing the productivity of L-glutamic acid of microorganisms in order to respond to further increase of demand of L-glutamic acid in future.
On the other hand, a gene encoding pyruvate carboxylase of Corynebacterium glutamicum (pyc gene) has already been cloned, and Corynebacterium glutamicum whose pyruvate carboxylase activity is enhanced by expression of the gene, and Corynebacterium glutamicum the same gene of which is inactivated have been known (Peters-Wendisch, P. G. et al., Microbiology, 144, 915-927 (1998)). However, these microorganisms were created as research materials used for the study of enzymes required for the growth of the microorganisms with glucose, and the relationship between pyruvate carboxylase activity and L-glutamic acid productivity has never been elucidated.